i
WHO Guidelines on Tissue Infectivity
Distribution in Transmissible Spongiform
Encephalopathies
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WHO Library Cataloguing-in-Publication Data
WHO guidelines on tissue infectivity distribution in transmissible spongiform encephalopathies.
1.Creutzfeldt-Jakob syndrome – pathogenicity. 2.Creutzfeldt-Jakob syndrome – diagnosis. 3.Encephalopathy, Bovine spongiform – transmission. 4.Encephalopathy, Bovine spongiform – pathogenicity. 5.Prion diseases. 6.Blood transfu- sion. 7.Pharmaceutical preparations. 8.Biological products. 9.Guidelines. I.World Health Organization.
ISBN 92 4 154701 4 (NLM classifi cation: WL 300) ISBN 978 92 4 154701 7
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EXECUTIVE SUMMARY v
INTRODUCTION 1
REVIEW OF SCIENTIFIC DEVELOPMENTS 2
Epidemiology, clinical features and diagnostic criteria of Creutzfeldt-Jakob disease (CJD) 2 Bovine spongiform encephalopathy (BSE) and scrapie 3
Diagnosis 4
Risk of transmitting Creutzfeldt-Jakob disease (CJD) and variant CJD (vCJD)
by human blood and blood products 5
RECOMMENDATIONS 7
Tissue infectivity 7
Measures to minimize risks to humans from biological and pharmaceutical
products in which ruminant materials are used during manufacture 7
Source of starting materials 8
Tissue removal and processing 8
Production systems 9
Vaccines 9
Recombinant DNA Products 10
Other medicinal products 10
Measures to minimize risks to humans from human-derived materials 11 The risk of transmitting variant CJD (vCJD) by blood and blood products 11
Risk assessment 12
Risk-reducing measures 13
Product retrieval and market withdrawal 13
Donor deferral 13
Deferral of transfusion recipients as blood donors 14
Plasma products 14
Appropriate blood usage 14
Other measures 14
Future developments 15
The risk of transmitting vCJD by human cells, tissue and tissue-derived products 15
CONCLUSIONS 16
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ANNEXES WITH REFERENCES
Annex 1: Major categories of infectivity (human TSEs, cattle, sheep, goats) 17
Annex 2: Summaries of scientifi c presentations 26
Epidemiology 26
Epidemiology update on human TSE diseases 26 Epidemiology update on animal TSE diseases 26 Update on vCJD prevalence estimated from tonsil and appendix screening 27 Progress in Detection and Quantitation of Infectivity 28
Bioassays 28
Primates 28
Transgenic mice 29
Bank voles 29
In vitro assays/cell cultures 29
Cautionary artifacts in detection of PrPTSE 30
Tissue or Body Fluid Infectivity 30
TSE infectivity in muscles and peripheral nervous system 30
CJD patients 30
Sheep with scrapie 30
BSE, including an update and methodology 31
Experimental models of TSE diseases 33
Tissue infectivity in urine 34
TSE blood infectivity 34
Transfusion transmission of vCJD 34
Scrapie or BSE-affected sheep 35
Experimental models of TSE diseases 36
Evaluation of TSE Blood Transmission Risk 37
Blood screening tests 37
Approach to validation of tests: strategies for development of reference materials 38
Evaluation of TSE removal procedures 40
Labile blood products 40
Plasma derivatives 41
Decontamination: new procedures 42
Assessment of the risk of transmission of vCJD via blood, blood components
or plasma derived products 42
ACKNOWLEDGEMENTS 45
v
EXECUTIVE SUMMARY
A variant form of Creutzfeldt-Jakob Disease (vCJD), a fatal brain disease of humans, fi rst appeared in the mid-1990s, as a result of the bovine spongiform encephalopathy (BSE or “mad-cow” disease) epidemic in the United Kingdom. Since fi rst reported in 1996, up to June 2006, 161 cases of vCJD have occurred in the United Kingdom, 17 in France, four in Ireland, two cases in USA and the Netherlands and single cases in Canada, Italy, Japan, Portugal, Saudi Arabia and Spain. Cases of BSE and vCJD have been decreasing in the United Kingdom in recent years, but both diseases have appeared in other countries.
Until recently, all vCJD cases were attributed to consumption of beef products con- taminated with the infectious agent of BSE. Since December 2003, three individuals have been identifi ed with vCJD infections probably acquired from blood transfusions – two with typical vCJD symptoms and the other with pre-clinical vCJD involving spleen and lymph nodes but not brain. Th e fact that the three vCJD infections followed transfusions from clinically healthy persons who became ill more than a year after donating blood implies that other blood donors who might currently be incubating the disease would also be potential sources of infection for recipients. Th e possible extent of future blood-borne spread of vCJD infections is still unknown.
Th e identifi cation of these cases has intensifi ed the concern about possible unmapped ways in which the disease might spread. Except for the three transfusion-transmitted infections, no cases of vCJD have been linked to any medicinal product to date, and guidelines have been developed by the World Health Organization (WHO) and other authorities to minimize the risk.
A Consultation held at the WHO in September 2005 brought together experts and regu- lators from around the world to revise existing WHO Guidelines on Transmissible Spongiform Encephalopathies (TSEs) in relation to Biological and Pharmaceutical Products (2003) which recommended ways to prevent potential transmission of vCJD through human blood and blood products, as well as through medicinal products prepared with bovine derived materials. Th e primary aim of the Consultation was to provide evidence-based information to national regulatory authorities of WHO Member States – especially to those where BSE has not yet been reported and where surveillance systems for BSE and vCJD may not be in place. Th e information is intended to assist them in conducting risk assessments and selecting measures to reduce the risk of transmitting vCJD by human blood and blood products and other medicinal products of biological origin, collectively called biologicals.
Th e WHO Guidelines of 2003 encouraged authorities to consider introducing precau- tionary measures to minimize the then-theoretical risk of transmitting vCJD by blood and blood products while not compromising an adequate supply. While acknowledging that transfusion- transmitted vCJD is no longer just a theoretical possibility, experts in the 2005 WHO Consulta- tion again advised authorities to assess the vCJD risk in the context of their own national situations, weighing the potential benefi ts of adopting precautionary policies to reduce that risk against the impact that those policies might have on the supply of blood.
Earlier WHO Consultations repeatedly stressed that, when feasible, tissues or body fl uids of ruminant origin should be avoided in the preparation of biological and pharmaceutical products.
When bovine materials must be used, they should be obtained from sources assessed to have negli- gible risk from the infectious agent of BSE. Most bovine tissues, including bovine muscle, used to manufacture biologicals, if carefully selected by taking into account the geographical distribution of BSE and collected according to guidelines, have little risk of contamination with BSE agent.
Recent fi ndings of disease-associated proteins in muscles of sheep with scrapie (a disease similar to BSE but not known to infect humans) and the recognition of BSE itself in a goat, reinforce the need for manufacturers of biologicals to maintain the precautionary safety measures recommended in the previous WHO guidelines. Ruminant blood and blood derivatives, such as fetal calf serum
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in cell cultures media and bovine serum albumin stabilizers, are also used to prepare biologicals.
Bovine blood has not been identifi ed as a source of infection, and properly collected fetal bovine serum has a negligible risk. However, the blood of sheep with experimental BSE or natural scrapie can be infectious and, because scrapie and BSE agents behave similarly in sheep and goats, the blood of small ruminants should either be avoided in preparing biologicals or selected very carefully from sources known to be free of TSEs.
Th ere is a continuing need to ensure that all national regulatory authorities with limited resources have ready access to reliable and up-to-date information when assessing TSE risks and evaluating product safety. Th at information includes guidance to help minimize or eliminate the risk for transmitting BSE and vCJD to humans via biologicals and other medicinal products.
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1Because of increasing complexity in terminology for various forms of the prion protein (PrP), the Consultation used the term PrPTSE for all abnormal misfolded PrP associated with TSEs.
2Th e third case was reported in February 2006 (http://www.eurosurveillance.org/ew)
INTRODUCTION
Th e WHO Guidelines on Transmissible Spongiform Encephalopathies in relation to Biological and Pharmaceutical Products, published in 2003, (http://www.who.int/bloodproducts/TSE/en) defi ned major categories of TSE infectivity, both in human and animal tissues, and have formed the basis of worldwide regulations for biological and pharmaceutical products. Preventive measures to minimize the risks of transmission to humans from both animal and human derived materials were also included in those Guidelines. New scientifi c information regarding the distribution of infectivity in various tissues from diff erent species aff ected with a transmissible spongiform encephalopathy (TSE or prion disease) has emerged during the last three years. Some of these fi ndings have challenged the current understanding of the tissue distribution of the pathological misfolded prion protein (PrPTSE)1 and tissue infectivity. An example of these is the fi nding of PrPTSE in sheep muscle. Th is is the fi rst evidence of PrPTSE in muscle from an animal species that enters the human food chain.
Also of concern has been the identifi cation of three cases of probable transfusion-transmitted vCJD2, suggesting that secondary transmission of vCJD from human to human has occurred. Th e demonstration of vCJD transmission via blood creates a special concern for those countries that have no human TSE surveillance system in place. Blood donors subsequently diagnosed as suff ering from vCJD have been identifi ed in the UK, France, Ireland, Saudi Arabia and Spain. It is clear that the blood of donors incubating vCJD might contribute to an unrecognized spread of the disease, especially in countries where surveillance and reporting systems are not established. Th is new concern about human-to-human transmission should not distract from eff orts to estimate and reduce the well established risk of food-borne BSE agent leading to vCJD cases. Each country should evaluate the risk of vCJD from both sources.
Th e development of reliable diagnostic procedures to detect asymptomatic subjects during the long periods of incubation of CJD and vCJD is of vital importance. However, test methods must be appropriately validated, and validation requires that appropriate blood reference materials be developed, characterized and made available both to qualifi ed test developers and to regulatory authorities.
In order to review and summarize the latest data on the epidemiology of vCJD, and on the detection of the infectious agents in blood, as well as the distribution of infectivity (and the abnormal prion protein associated with infectivity) in other tissues or body fl uids of relevant species with TSEs, a Consultation of international experts was convened at WHO in Geneva on 14-16 September 2005.
Th e analysis of the information presented provided scientifi c evidence to support the updating of the WHO Recommendations published in 2003. Experts at the Consultation were also asked to consider whether additional measures should be recommended to maintain the safety of blood and blood products, vaccines and other medicinal products with respect to TSEs.
Policies developed to reduce the risks resulting from the hazard of vCJD and its presumptive human-to-human transmission are based on three main factors: (a) an unknown number of individu- als have been infected with the BSE agent; (b) the pathological misfolded prion protein found in TSEs – here designated as PrPTSE – and infectivity, detected by bioassay, are present in some peripheral tissues of patients who died of vCJD; (c) blood of rodents, sheep and possibly non-human primates infected with TSE agents has transmitted disease experimentally and, three persons in the UK have very probably been infected with the vCJD agent from the blood of asymptomatic donors who later died of vCJD. Th ere is increasing concern that, without appropriate controls, blood products and other pharmaceuticals manufactured using bovine-derived or human-derived materials might spread the agent of vCJD worldwide, even to countries where BSE has not been reported. Th ere is, therefore, a need to ensure that regulatory authorities worldwide have reliable information to assess the risk of human exposure to the BSE/vCJD agent, so that steps can be taken to prevent the transmission of TSE to humans via biologicals and other pharmaceutical products.
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REVIEW OF SCIENTIFIC DEVELOPMENTS
Changes in the distribution and size of the BSE epidemic in Europe and elsewhere continue to be observed. Th e distribution of infectivity in tissues and body fl uids in sporadic CJD (sCJD), vCJD, BSE and scrapie has been better established and methods for detecting the PrPTSE in diff erent tissues have improved. Summaries of the information reported by the experts in the Consultation together with the relevant scientifi c references are provided in Annex 2.
Epidemiology, clinical features and diagnostic criteria of Creutzfeldt-Jakob disease (CJD)
CJD is a rare and fatal human neurodegenerative condition. Like other TSEs, CJD is experi- mentally transmissible to animals, and a characteristic spongiform change is seen on microscopic examination of the brain. Epidemiological studies indicate a worldwide occurrence of sporadic disease of approximately 1-2 cases per million people per year. Globally, over 80% of cases of CJD occur as a sporadic disease (sCJD). Familial, iatrogenic, and variant forms of CJD show much lower and variable incidence in diff erent countries. Most cases of vCJD have been found in the UK.
Th e origin of sCJD remains unknown despite extensive study. In particular, there is no evidence of a causal link with scrapie, a naturally occurring TSE of sheep and goats, or with BSE.
Most sCJD cases occur in persons between the ages of 60 and 80 years with an average age at death of about 67 years. Th e typical patient with sCJD develops a rapidly progressive dementia associated with multifocal neurological signs, ataxia, and myoclonus. Although, in the correct clinical context, a characteristic EEG recording and/or the detection of 14-3-3 protein in the cerebrospinal fl uid are considered diagnostic, confi rmation of the diagnosis of CJD still relies on neuropathological examination. Th e clinical and histopathological features of sCJD are variable and are infl uenced by a naturally occurring polymorphism at codon 129 of the gene encoding the prion protein (PRNP gene). A novel test based upon the detection of PrPTSE in the nasal olfactory mucosa may improve the diagnosis of sCJD.
Familial CJD, also experimentally transmissible, is expressed as an autosomal dominant trait associated with one of several abnormalities of the PRNP gene in diff erent aff ected kindreds.
Gerstmann-Sträussler-Scheinker syndrome (GSS) and fatal familial insomnia (FFI), similarly inherited neurodegenerative disorders linked to mutations in the PRNP gene, have also been transmitted to animals.
Th ere have now been at least 362 recognized cases1 of iatrogenic CJD following use of the following products: contaminated human-pituitary-derived growth hormone (180 cases) and gonadotropin (four cases), human dura mater allografts (168 cases), corneal transplants (at least three probable or possible cases), neurosurgical instruments (four or possibly fi ve cases) and a stereotactic cortical-probe EEG electrode (two cases). Except for the three transfusion-transmit- ted vCJD infections described above, no new class of products causing iatrogenic CJD has been identifi ed during the last nine years, although the number of cases resulting from past exposures to known products continues to increase.
Since fi rst reported in 1996, up to June 2006, there have been 161 cases of vCJD in the United Kingdom, 17 in France, four in Ireland, two in the USA and in the Netherlands and single cases in Canada, Italy, Japan, Portugal, Saudi Arabia and Spain. Cases of BSE and vCJD have been decreasing in the United Kingdom in recent years, but both diseases have appeared in other countries and the notifi cation rate for new cases of vCJD has increased in France during the past two years. Th e US, Canadian and two of the Irish patients had spent several years in the UK
1Th e numbers of all iatrogenic cases were kindly provided by Dr Paul Brown (October 2005)
3 between 1980 and 1996, and were probably exposed to the BSE agent there, while the Italian, the
Dutch cases and 16 of the 17 French cases had no history of signifi cant travel outside their home countries. Th e median age at onset of vCJD is 26 years (range 12-74 years) with a median duration of illness of 14 months (range six to 40 months). Although the defi nitive diagnosis of vCJD requires neuropathological examination, clinical and laboratory criteria have been established to diagnose probable vCJD in living patients. All clinically manifest vCJD cases (191 worldwide) tested were homozygous for the allele encoding methionine at codon 129 of the PRNP gene. A distinctive feature of vCJD—in contrast to sCJD—is the frequent occurrence of PrPTSE in lymphoid tissues (tonsil, spleen, lymph node, and appendix). In three patients who underwent appendectomy before onset of vCJD, two had immunoreactive PrPTSE in lymphoid follicles of the appendix—one eight months and the other two years prior to death. One appendix was negative for PrPTSE nine years prior to death. An anonymous survey of surgically removed tonsils and appendices in the UK revealed three out of 12,674 cases which stained positively for PrPTSE. All three were appendices.
Genetic studies on DNA extracted from two of the three positive appendices found that they were valine homozygous at codon 129 in the PRNP gene, unlike any of the clinical case of vCJD encountered so far.
Th e causal link between vCJD and BSE is based on epidemiological, biochemical and transmission studies. A Joint Technical Consultation on BSE 2001 convened by the WHO, the Food and Agriculture Organization (FAO) and the Offi ce International des Epizooties (OIE, the World Organization for Animal Health) reached a scientifi c consensus that consumption of beef products contaminated with the BSE agent is the main avenue of exposure (http://www.who.int/
zoonoses/diseases/en); this conclusion is still generally accepted. Bovines, bovine products and by- products potentially carrying the BSE agent have been traded worldwide, giving this risk a global dimension. Epidemiological analysis does not indicate that most medicinal products, including plasma-derived products, or occupational exposures have been sources of infection in vCJD cases identifi ed to date. Th ree cases of vCJD infection presumptively transmitted by transfusion of red blood cell concentrates have been reported.
Bovine spongiform encephalopathy (BSE) and scrapie
BSE was fi rst identifi ed in British cattle in November 1986. Current evidence suggests that the disease originated from the use of feed supplements containing meat-and-bone meal contaminated with a TSE agent (probably from scrapie-infected carcasses). In the UK, 184,370 confi rmed cases of BSE had been reported by 31 December 2005 (http://www.oie.int/eng/info/en_esbru.htm).
Smaller outbreaks have been reported in native-born cattle in most other Western and Central European countries and in Canada, Israel, Japan, and the USA. Outside the UK, 5,428 confi rmed cases had been reported to the OIE or the EC by 31 December 2005. Most recent cases were recognized either in cattle at increased risk for BSE (fallen stock and emergency slaughter cattle over 24 months old) or in clinically unremarkable slaughter cattle over 30 months old identifi ed by statutory or other “rapid” testing (see Diagnosis) of brainstem tissue for PrPTSE. Such tests were introduced in Switzerland in 1999 and in the European Union in 2001. Active testing of all cattle was instituted in Japan after 2001 and in high-risk animals in Canada and the USA after 2003. Such targeted active surveillance in Europe has resulted in the better detection of infected animals during the pre-clinical and clinical stages of illness. In the UK, the incidence of BSE has continued to decline since 1992-1993, consequent to a statutory ruminant feed ban introduced in 1988 and reinforced in 1996. Th is decline is consistent with the hypothesis that BSE cases arose by infection from contaminated feed. Although epidemics of BSE in other European countries were recognized more recently than that in the UK, most are also in decline, and, so far, no single country except the UK and Ireland has reported more than 1500 cases.
In naturally aff ected cattle, BSE infectivity, detected by assay in mice, has been demon- strated only in the brain, spinal cord and retina. Assays of infectivity in cattle have also detected infectivity in a pool of nictitating membranes but not in pools of lymph nodes or spleen. Recently, infectivity was detected in some peripheral nerves and a solitary muscle, of a single case of BSE in a
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German cow, assayed using highly sensitive transgenic mice over-expressing bovine PrP, suggesting a very low titre of infectivity.
In cattle experimentally exposed by the oral route, BSE infectivity has been detected by mouse assay in the distal ileum through much of the disease course from six months post exposure and in the central nervous system (CNS) and in sensory ganglia of the peripheral nervous system from late in the incubation period. Infectivity has also been detected in sternal bone marrow in cattle experimentally exposed to BSE agent by the oral route, but only at a single time point during clinical illness. Assays in cattle of selected tissues from this same initial sequential time point oral exposure study confi rmed infectivity in distal ileum (from six through 18 months after exposure and during clinical disease) and in the CNS at the earliest time post-exposure detected by the mouse assay but not before. Infectivity has also been found in palatine tonsil, at a single time point in incubation, but this was detected only by assay in cattle and not by the mouse assay. A wide range of other tissues, including most lymphoreticular tissues, from cattle with BSE—both naturally acquired and experimentally induced—and from cattle in the incubation period after experimental exposure, contained no infectivity detectable by conventional mouse bioassays or ongoing parallel bioassays in cattle. Bioassays of bovine tissues injected into transgenic mice over-expressing bovine PrP, presumably reducing the PrP-associated species barrier, also support a conclusion that there is a limited distribution of BSE infectivity in bovine tissues.
BSE has been experimentally transmitted via the oral route to sheep and goats, and there is recent evidence that one goat has been naturally infected. Concern over the possibility of BSE in small ruminants led to increased eff orts at active and passive surveillance of scrapie in the EU, based on the observation that experimental BSE in sheep and goats resembles scrapie.
Recently, infectivity was found in blood of sheep with natural scrapie and in blood of sheep with experimental BSE during both the incubation period and clinical illness.
Diagnosis
Th e accumulation of PrPTSE occurs only in TSEs, so its detection can serve as a surrogate for detection of infectivity in biological samples. After experimental inoculation of rodents with TSE agents, PrPTSE, like infectivity, is usually detectable in the CNS weeks before the appearance of overt disease, and its level increases during clinical illness. While the increase in PrPTSE generally parallels that of infectivity, the precise relationship between PrPTSE and infectivity is unclear. For example, under specifi c experimental conditions, the brains of some TSE-aff ected rodents may be infectious by bioassay while PrPTSE remains undetected. From the perspective of pre-clinical diagnosis, both the sensitivity of diagnostic methods and procedures to concentrate PrPTSE are crucial, because the amount of PrPTSE outside the CNS is likely to be small, particularly in circu- lating blood. PrPTSE can be concentrated by physicochemical precipitation, affi nity precipitation techniques or affi nity chromatography. Moreover, it has been reported that the amount of PrPTSE in dilute solutions can be increased considerably by the “protein misfolding cyclic amplifi ca- tion” (PMCA) technique, potentially allowing improved detection of extremely small amounts of infected material. In recent reports, the test developer described an improved PMCA that detected PrPTSE in the blood of most hamsters with scrapie and not in the blood of uninfected hamsters.
In addition to blood, other readily accessible tissues might off er the possibility for diagnosis of clinical TSE. Tonsil biopsy has been used to diagnose vCJD in a minority of patients after the onset of clinical signs and symptoms. Also, a study that demonstrated PrPTSE and infectivity in the skeletal muscle of mice experimentally infected with laboratory strains of TSE has been at least partially confi rmed by the detection of PrPTSE in skeletal muscles of small ruminants with TSEs and humans with both sporadic CJD and vCJD. Th ese fi ndings are under intense study by a number of laboratories.
Among immunological methods for PrPTSE detection, immunoblotting (Western blotting) is the most thoroughly characterized and widely used. Immunoblotting off ers the advantage of recognizing diff erent forms of PrPTSE through the analysis of the molecular mass, shift in electro- phoretic mobility after digestion with proteinase K (PK), relative abundance of di-, mono- and non-glycosylated bands, and binding with a variety of epitope-specifi c monoclonal antibodies.
5 Th e size of PK-treated fully deglycosylated PrP and relative abundance of di-, mono-and non-
glycosylated bands characterize the PrP type, a kind of PrP signature that varies among diff erent forms of TSE. PrPTSE typing has been proposed for distinguishing various forms of TSE (e.g., scrapie from BSE and sCJD from vCJD) and for improving the classifi cation of human TSEs.
ELISA and immunoblotting methods are commercially available in Europe, Canada and the USA as ready-to-use kits for postmortem animal diagnosis of TSEs; several of those tests (so-called “rapid” tests) have been validated in a study by the European Commission (EC) as screening tests for BSE in slaughtered cattle. EC-approved immunoassays have detected PrPTSE in the brains of BSE-infected cattle at least three months before onset of clinical illness. However, no immunological method has yet been validated to be suffi ciently sensitive to detect PrPTSE in the blood of infected animals or humans, though promising initial results have been reported by several groups of investigators and were presented to the Consultation (see Annex 2). Some immunoassays have been claimed to detect PrPTSE in samples containing less than 1 LD50 of BSE infectivity as measured by bioassay in transgenic mice expressing bovine PrP, though attempts to confi rm the claims independently have not been successful.
Risk of transmitting Creutzfeldt-Jakob disease (CJD) and variant CJD (vCJD) by human blood and blood products
Since the last WHO Consultation on this issue in 2003, new evidence relevant to risk assessments for the transmission of vCJD by human blood has accrued. Salient information is summarized here:
(a) It has been known for more than 20 years that the blood of rodents experimentally infected with agents of several TSEs contains infectivity. Most recently, infectiv- ity has been found in the blood of mice experimentally infected with the agent of vCJD.
(b) Th ere is convincing evidence that both scrapie and BSE can be transmitted from sheep to sheep by blood transfusions with either whole blood or buff y coat. Transfusions of blood from animals in the incubation period and clinical phase of illness have transmitted disease. Transfusion appears to be a relatively effi cient mechanism for transmitting infection from sheep to sheep.
(c) Epidemiological evidence indicates that vCJD infection has been transmitted to three recipi- ents of blood transfusion. Th ese three infected recipients demonstrate that blood contained infectivity during the latter part of the incubation period of vCJD, from 18 months to 3.5 years before the donors showed signs of neurological illness. Th e fi nding of three transfusion- transmitted vCJD cases among a relatively small number of persons transfused with blood components from vCJD donors, only about 18 of whom survived for more than fi ve years, suggests that the transfusion of a human blood component has transmitted vCJD effi ciently, an observation consistent with experimental animal studies. Th e fi rst recipient developed vCJD 6.5 years after transfusion—considerably shorter than the probable minimum incuba- tion periods of presumed food-borne cases of vCJD. Th e second recipient died without signs of neurological disease fi ve years after transfusion but already had detectable PrPTSE in spleen and lymph nodes, though not in appendix, tonsil, or brain. Th e third patient, who developed clinically typical probable vCJD almost eight years after transfusion of red cells from a diff er- ent donor, was still alive at the time of the Consultation in 2005 but has subsequently died.
(d) To date, all cases of vCJD tested have been in persons homozygous for methionine at codon 129 of the prion-protein-encoding gene (PRNP gene). However, the transfusion recipi- ent with pre-clinical or sub-clinical vCJD infection was heterozygous, having methionine and valine at PRNP codon 129, indicating that vCJD infection can occur in persons of this genotype, as can other forms of CJD. Furthermore, a study of anonymous tonsil and appendix specimens in the UK identifi ed three instances in which appendix samples stained positively for PrPTSE using immunohistochemical techniques—although the staining pat- tern was described as being slightly diff erent from that in lymphoid tissues of known vCJD cases. Genetic studies on DNA extracted from two of these three appendices found them to
6
be valine homozygous at codon 129 in the PRNP gene; no DNA was extractable from the other specimen. Taken together, these fi ndings suggest that, if exposed to a suffi cient dose, most people are probably susceptible to infection with the BSE agent. In the UK, approxi- mately 30% of Caucasian populations are homozygous for methionine at codon 129 of the PRNP gene, about 50% are heterozygous for methionine/valine, and the rest are homozygous for valine. Th e PRNP codon-129-valine allele is rarely found in East Asian populations.
(e) Th e same tonsil-appendix survey results also suggest that a substantial number of individuals in the UK might be incubating vCJD—a mathematical analysis predicting that as many as 5,000 individuals in the total UK population (a rate of 237/million) might be infected. Some proportion of healthy individuals with sub-clinical or pre-clinical vCJD would presumably be blood donors.
Th e possible prevalence of asymptomatic vCJD infections in other countries is not known.
(f) A probabilistic risk assessment model concluded that transmissions of infection by blood transfu- sion had a potential to increase the eventual size and duration of the current vCJD outbreak in the UK signifi cantly. Deferral of transfusion recipients as blood donors was implemented in the UK in 2005; this step is anticipated to reduce substantially the risk of recycling vCJD infections. Th e same measure has been in place in France since 1998 and more recently in several other European countries such as Ireland, the Netherlands and Switzerland. In some other countries, like Canada, Australia, Italy and the US, blood donors previously transfused in the UK have been deferred.
(g) A risk assessment estimated that some derivatives prepared from pools of UK plasma were likely to have included one or more donations from persons incubating vCJD. Th at poses a small and uncertain risk of transmitting infection to some recipients of the prod- ucts. Th e relative risk depends on the type of plasma product, the specifi c manufactur- ing process used and the year of production. Th e UK has no longer fractionated plasma of UK residents since 1999 and has imported all plasma for manufacture—most from the USA and some from Germany. Assessments suggested that vCJD risk from deriva- tives of plasma collected and manufactured in other countries was low or negligible.
(h) A few vCJD cases in the UK were in people previously transfused with blood compo- nents from donors not diagnosed with vCJD, and a risk assessment concluded that some of these recipients might conceivably have been infected through the transfusion. For two cases, the time between blood transfusion and onset of vCJD was so short that transfu- sion transmission seems highly improbable. Authorities have not concluded that any of the donors involved is sub-clinically infected with vCJD agent. Nevertheless, the impli- cated donors were informed that, as a precaution, they should no longer donate blood.
In conclusion, it is probable that vCJD has been transmitted through blood transfusion, with important implications for public health. Several vCJD cases have occurred outside the UK in persons who previously donated blood (in France, Ireland, Saudi Arabia and Spain). Authorities in those countries are aware of the potential risk. To date there is no evidence that vCJD has been transmitted by human plasma derivatives, in spite of intensive use of some products manufactured from plasma of UK donors during and after peak years of the UK BSE outbreak. But the incuba- tion periods of TSEs can be very long, and possible transmission of vCJD by plasma derivatives, while not recognized to date, cannot be confi dently excluded yet. Cumulative epidemiological evidence, including follow-up studies in Europe and the USA of more than a hundred long-term survivors transfused with blood components from donors who later developed other forms of CJD, suggests that the infectious agents associated with sporadic, familial and iatrogenic CJD have probably not been transmitted through blood or blood products, at least not with a frequency detectable by epidemiological surveys.
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RECOMMENDATIONS
Tissue infectivity
Th e foundation of any attempt to construct a rational analysis of TSE risk for biological and other pharmaceutical products must begin with an evaluation of infectivity in the human or animal tissues from which these products are derived. Although straightforward in principle, the task is complicated by diff erences in the timing of fi rst appearance and fi nal tissue distribution of infectivity in diff erent species and TSEs, by diff erences in the sensitivity of bioassay methods, and by incomplete data about infectivity levels in various tissues of interest. Tables IA, IB and IC in Annex 1 summarize current data about the distribution of infectivity and PrPTSE in humans with vCJD and other human forms of TSE, in cattle with BSE, and in sheep and goats with scrapie.
In general, it can be said that, paradoxically, whereas both infectivity and PrPTSE in cattle with BSE have a more limited distribution in tissues than in any other animal or human form of TSE, PrPTSE and, to some extent, infectivity have a wider distribution in tissues of humans with vCJD than in other forms of CJD. In using the tables, it is important to note that two classes of material were intentionally excluded: (1) materials like bile of ruminants and humans that have never been studied, and (2) highly processed chemically pure reagents like tallow derivatives and bovine bone gelatin produced by the alkaline process that have been studied and found to pose a negligible risk if any for transmitting infection.
Several new methods attempting to detect PrPTSE using novel techniques (see Diagnosis and Annex 2), if successfully developed, might eventually off er suffi cient sensitivity to demonstrate amounts of agent below the level of detection of currently validated tests. It has been speculated that such methods might fi nd small amounts of agent in some tissues currently thought to be free of infectivity. It remains unknown whether tissues containing such very small amounts of infectious material would transmit infection to humans.
Measures to minimize risks to humans from biological and pharmaceutical products in which ruminant
materials are used during manufacture
Bovine materials are commonly used in the manufacture of many biological and pharmaceutical products. Th e use of ovine and caprine materials—except for the occasional use of sheep and goat antisera and the milk of transgenic goats—is relatively uncommon. On the basis of current scientifi c knowledge about the agents causing BSE and other animal TSEs, an ideal strategy would be to avoid altogether the use of ruminant (especially bovine) materials in the manufacture of any biological or pharmaceutical product, as well as the use of materials from other animal species in which TSEs naturally occur. In practice, this is often not feasible, in which case measures should be taken to minimize the risk of incorporating an infectious TSE agent. A risk assessment should be performed for the fi nal product. Th e risk assessment should take into account three general factors: source of starting materials, manufacturing process and clinical use of the fi nal product.
Th e sources of starting materials used as active substances, excipients or manufacturing reagents and their potential infectivity are most important. An assessment should include both the geographic source (country), history of the source animals (age, feeding, traceability), and details of the actual tissues used (risk of intrinsic infectivity in the tissue and risk of cross-contamination with higher-risk materials during slaughter or processing of tissue). It is generally acknowledged that source tissues processed using validated methods are likely to pose a lower risk than unprocessed or minimally processed source tissues. Th is is especially true when the processing conditions have a validated robust capacity to inactivate or remove TSE agents, such as those used in the manu- facture of amino acids, tallow derivatives, and gelatin. For example, gelatin prepared from bovine bones—excluding skulls and vertebrae—subjected to a series of partially eff ective but additive processes such as hot-water washing under pressure, demineralization with acid, alkaline hydrolysis
8
and, especially, terminal ultra-high-temperature fl ash heat sterilization of the fi nal product poses little risk.
An assessment of risk should consider the ability of a manufacturing process to inactivate infectivity or to remove infectivity by partition, as well as the possibility of cross contamina- tion—both when starting materials are collected and processed and during manufacture of the medicinal products themselves. Th e production processes for preparing master and working viral and bacterial seeds as well as cell banks and other materials used in the manufacture of vaccines should not be neglected.
In considering the clinical use of the product in a risk assessment, both the route of administration and amounts of product likely to be used in a single course of therapy, in a year or even in a lifetime should be taken into account. A thorough TSE risk assessment contributes to the overall risk-benefi t analyses for biological and other pharmaceutical products.
Source of starting materials
Careful selection of the source of ruminant starting materials used to manufacture active sub- stances, excipients and in-process reagents is an important consideration in the TSE risk assess- ment. Th e most satisfactory source of materials is from countries where the risk of BSE in cattle is low and adequately controlled. Countries are encouraged to assess geographic BSE risk, and the OIE off ers guidance in that process through the latest edition of the OIE Terrestrial Animal Health Code in its chapter on BSE (http://www.oie.int/eng/normes/mcode/A_summary.htm).
OIE currently suggests that countries assign themselves to one of three categories of BSE risk:
negligible, controlled or undetermined. National regulatory authorities may use their own more refi ned estimates of BSE risk for purposes of import controls. For the EU, the Geographical BSE Risk categorization—a standard qualitative approach to assess the risk of BSE for cattle in countries submitting requests to trade—has been expanded and updated continuously, originally by the Scientifi c Steering Committee of the EC (Health and Consumer Protection Directorate General) and subsequently by the European Food Safety Authority (EFSA, at http://www.efsa.eu.int/sci- ence/tse_assessments/catindex_en.html). Th e Geographical BSE Risk categorization recognizes the important fact that if the prevalence of BSE in a country varies over time so too will the result of its risk assessment.
Th e use of ruminant source materials from countries with an undetermined risk of BSE is usually not acceptable. However, even in those countries, the collection of source materials for the manufacture of specifi c products from well-monitored herds might sometimes be allowed, particularly if they are from tissues appearing in Table IC and they are collected from healthy cattle in a safe manner. Th is should be done only if evidence is provided that the herds have had no cases of BSE, and have implemented an active BSE surveillance program. In addition, these ruminants should have never been fed meat-and-bone meal nor any other mammalian-derived proteins prohibited by national authorities (including some processed animal proteins) and have a fully documented breeding history, including introduction of new genetic material only following OIE guidelines for international trade in bovine semen and embryos.
Th e age of animals from which tissues or fl uids are collected as starting materials should also be taken into account. Studies on infectivity of tissues collected during the incubation period of experimental BSE in cattle showed that tissues of younger animals generally contained less infectivity than did those of older animals. However, some bovine tissues, like ileum and tonsil, contained infectivity at an early stage following oral exposure to the BSE agent. Such tissues may be considered safe only where the risk of BSE in cattle is negligible or where epidemics of BSE have been in continuous decline and as long as rigorous high-quality surveillance and BSE control procedures remain implemented.
Tissue removal and processing
Potential TSE risks might be infl uenced by circumstances under which tissues are removed. For example, both penetrative and some non-penetrative techniques for stunning cattle before terminal exsanguination can embolize brain tissue into the general circulation and increase the risk that
9 tissues containing little or no intrinsic infectivity (e.g., lung) might become contaminated with
high-risk tissue. Stunning devices that inject air into the cranial cavity and the process known as
“pithing” are especially dangerous in this regard. Methods to prevent cross-contamination of the carcass with brain from skull wounds and from neural tissue exposed during decapitation should be in place. Sawing of the vertebral column introduces another possibility for contaminating intrinsically low-risk tissue with spinal cord. Collecting selected tissues before the carcass is split greatly reduces that risk. Body fl uids should be collected with minimal damage to tissue, and cel- lular components should be removed. Blood of young animals with known provenance is less likely to pose the risk of cross-contamination. Fetal blood should be collected without contamination from other maternal or fetal tissues, including placenta, and whenever possible, by taking blood by cardiac puncture with single-use instruments using closed collection systems for all materials.
When potential cross-contamination of a source tissue with a tissue of higher risk cannot be reasonably excluded, a higher risk of infection must be assumed for purposes of risk evaluation. In short, particular care should be taken to avoid any kind of contact between collected materials and higher-risk materials. Some national authorities have defi ned certain bovine materials of greatest concern as “specifi ed-risk” materials and required removal of those materials from carcasses.
Facilities that provide starting materials for medicinal products should have in place an appropriate quality system to document the process used and provide a record for each batch of starting material collected. Th ey should either have or work towards offi cial accreditation of the quality system. For example, the EU grants Certifi cates of Suitability for products complying with the EU Note for Guidance on TSE Risks, which stresses the need for a quality assurance system.
Procedures should also be in place to reduce the risks of adulteration of batches.
Th e sources and types of starting materials, while important, are not necessarily the only determinants of risk of potential TSE transmission. Some manufacturing processes—for example those used to produce bovine serum albumin and tallow derivatives—have a substantial demon- strated ability to eliminate infectivity that might be present in the starting material. Processes that inactivate infectivity or remove infectivity from starting materials augment the safety provided by appropriate sourcing. Manufacturers should consider including such procedures in their manu- facturing processes when possible. Claims that a production process contributes signifi cantly to the safety of a product should be validated.
Production systems
Vaccines
Production systems also aff ect the fi nal TSE risk assessment for vaccines. Many vaccines are prepared from organisms that cannot be treated with harsh methods of extraction or purifi cation without reducing or destroying their antigenicity. Additional diffi culties are inherent in the cell bank and seed lot systems employed in vaccines production. Concerns with respect to TSE may arise from the animals used for in vivo production or as a source of cells for production in vitro, from components of medium used in production, or from the cell banks or bacterial or viral seeds used to initiate production. Where vaccine strains are still grown in animals (such as vaccinia virus for smallpox vaccine), careful selection of source materials and, in some cases, postmortem testing of each production animal can greatly reduce the TSE risk. Some vaccines are produced in primary cell cultures, usually derived from species of animals not known to have TSEs. Such cultures are very unlikely to be infected or contaminated with TSE agents. Nevertheless, cells should be selected carefully, avoiding those known to replicate TSE agents. However, culture medium used to grow bacterial, yeast, mammalian or other cells in vitro may contain components of animal origin. A TSE risk assessment should be carried out for such production systems.
Th e most complex TSE issues are raised by banked eukaryotic or bacterial cells and viral vaccine seeds. Th e Consultation strongly emphasized that, by virtue of the level of characterization possible, the overall risk-benefi t assessment overwhelmingly favors the use of banked cells and the seed lot system for vaccine production. However, TSE risk assessments of banked cells and viral or bacterial vaccine seed stocks should take into account the possible carryover of any potentially infectious material from the seed into the fi nal product as a contaminant. Th ere is also a theoretical
10
possibility that production cells might be infected with a TSE agent, although none of the very few cell lines known to support replication of a TSE agent has been used to produce any vaccine.
When evaluating the possibility of potentially infectious material in a seed, not all relevant information may be available, since some seeds were derived long ago and often have a lifetime of decades. Tracing their origin and full production history may be impossible. Where information is incomplete, it is recommended that, when feasible, working seeds be replaced as a precaution- ary measure, taking into account the need to maintain adequate supplies of vaccines with public health benefi ts during the replacement. For existing products, master seed materials and original experimental preparations from which master seeds are derived (“pre-master” seeds) may not need replacement, since the biological phenotype of the vaccine depends on these materials, and they may be diffi cult or impossible to recreate. However, the goal of eventually replacing seeds with those of impeccable provenance for all reagents should not be abandoned. Th e origin of newly developed products should be documented as completely as possible, recognizing that this may also be diffi cult. For new products made using old seed lots, any existing risk assessment for the seed and history of prior use of the seed should be taken into account.
When deriving new vaccines with new cell banks or viral or bacterial seeds, developers should take into account all guidance on TSE risk in force at the time that laboratory work begins.
However, since development of new vaccines often takes years, complete information on TSE risk for older seed materials may not meet requirements in force later, when a candidate new product must be assessed for licensure. Th e principles on seed materials outlined in the paragraph above should apply to such cases.
Recombinant DNA products
Medicines produced by recombinant DNA technology use a cell banking system similar to that used for many vaccines. Similar considerations with respect to production media, carryover of contaminants and the theoretical possibility of infection of the cells therefore apply. Risk assess- ments should take the same approaches used for vaccines.
Other medicinal products
A number of bovine-derived materials are commonly used to manufacture both biological and pharmaceutical products. Th ese include milk and milk derivatives (like casein), meat extracts, bovine serum including fetal bovine serum, bovine bone gelatin, bile derivatives (deoxycholate, choline) and beef tallow derivatives (triglycerides, glycerol, sorbitol esters, polysorbates, other). Beef tallow prepared using specifi ed-risk materials should be avoided. Materials originating from non- bovine ruminants are less commonly used, although substantial amounts of mixed-species tallow may be produced from some rendering plants. Infectivity of experimental BSE is more widely distributed in tissues of small ruminants (see Annex 1) than in cattle, posing a special concern.
Milk and certain milk derivatives, such as lactose, are generally considered non-infec- tious, regardless of geographic origin, provided that the milk is from healthy cows fi t for human consumption and no other potentially infectious ruminant-derived materials were used in the manufacturing process. Rennet, derived from the abomasum of cattle—which has shown no detectable BSE infectivity (Annex 1, Table IB)—sometimes used during the manufacture of lactose is generally considered to pose no signifi cant risk, especially when derived from calves.
Extracts prepared from tissues like bovine muscle, in which infectivity was recently detected during the clinical phase of BSE in one cow (Annex 1, Table IB) are unlikely to present more than a negligible risk of TSE contamination, provided that the manufacturer has scrupulously complied with procedures designed to avoid cross contamination with specifi ed-risk materials during preparation of the source material. If assurances of compliance are not available, then it is recommended to source meat extracts from animals in countries where risk of BSE is remote.
Recently, using tests of increased sensitivity, infectivity and PrPTSE were detected in peripheral nerves of cattle and, PrPTSE in enteric plexus. An additional safeguard for bovine muscle used to prepare vaccines or other biologics might be to ensure removal of all visible nervous and lymphatic tissue from muscle before collection and to avoid using meat from the tongue or the head. A simpler approach would be to source meat for nutrient broths from young animals in countries where the risk of BSE is negligible.
11 As noted above, TSE infectivity has been detected in transfused blood from sheep with
natural scrapie and sheep experimentally infected with BSE. Transfusion experiments have not been conducted in cattle. Eff ects of blood clot formation on TSE infectivity in serum have not been established. Studies using small amounts of blood components or spleens of cattle with BSE assayed in mice and cattle injected by the most eff ective routes failed to detect infectivity.
A conservative regulatory approach would assume that bovine serum might potentially contain TSE infectivity—presumably in small amounts. Blood for the preparation of donor calf serum is most often collected from well controlled living animals, reducing the risk of cross-contamination of blood with higher-risk materials attendant to the stunning and slaughtering process. Th us the sourcing of bovine serum (country/herd/animal) combined with appropriate precautions to avoid cross-contamination during collection is important in the risk analysis. An additional safeguard might be to collect blood from impeccable sources and to store separated components for a period of time exceeding the mean incubation period of cattle—at least fi ve years. Such a strategy would increase confi dence in the safety of the material if no BSE had been detected in the country.
Gelatin may be extracted from the skin and/or bones of cattle and pigs. Skins of either species and bones of pigs are likely to have a negligible TSE risk, provided that contact with bovine specifi ed-risk materials is avoided. Gelatin from bovine bones originating anywhere except a country with negligible BSE risk should be produced by alkaline hydrolysis—including pressure washing with hot water, acid demineralization, and fi ltration, augmented, whenever possible, by an adequate fl ash ultra-high-temperature heat sterilization process—rather than by acid treatment alone. Bovine bones for gelatin should carefully exclude skulls and vertebral columns if obtained from countries other than those with negligible BSE risk. Compliance with these precautions provides assurance that gelatin used in the manufacture of medicinal products is unlikely to be contaminated. Amino acids derived from gelatin are further highly processed, so their risk may be even lower.
Materials derived from ruminant tallow (see above) and amino acids of ruminant origin, even if higher-risk tissues were not completely eliminated, are considered highly unlikely to remain contaminated by the time the fi nal reagents have been produced, so long as the reagents were prepared by processes of extraction and purifi cation at high temperature and pressure, and good manufacturing practices (GMP) were rigorously controlled. Safety is further assured when speci- fi ed-risk materials are excluded from starting materials, when raw materials are pressure cooked according to the OIE Code (particle size ≤ 50 mm, temperature > 133°C, pressure ≥ 3 bar, exposure time ≥ 20 min) and when proteins have been removed from tallow to meet OIE specifi cations.
Th e OIE Code chapter on BSE provides specifi c guidance on requirements for safe trading in commodities used to manufacture biologicals and other medicinal products.
Measures to minimize risks to humans from human- derived materials
The risk of transmitting vCJD by blood and blood products
Although the UK remains the country at greatest risk from past exposure to the BSE agent, vCJD has also been identifi ed in an increasing number of other countries in recent years. In most of those countries only single cases have been identifi ed. But the occurrence of even one vCJD case means that other people may be at risk of primary food-borne infection, either through consumption of imported BSE-contaminated beef products or by consumption of infected beef products during travel to the UK or to other increased-risk countries before eff ective BSE control measures were in place. Furthermore, some countries without recognized cases of vCJD might have unrecognized BSE in cattle or have unknowingly imported products contaminated with the BSE agent. Th ere- fore, it is likely that cases of vCJD will continue to occur, both in Europe and elsewhere. It would be prudent for national authorities to prepare, in advance, plans to reduce the risk of secondary transmission by blood components and plasma-derived products, even if BSE and vCJD have not been recognized in the country.
12
Risk assessment
Blood, blood components and plasma-derived products are essential in medical treatment and can be life-saving. An eff ective health care system should strive to ensure a consistent and adequate supply of these products. Some measures to reduce the risk of vCJD, for example by deferral of certain donors, might compromise supplies of blood and plasma. It is essential that any measures introduced in response to vCJD be proportionate to the risks, which vary from country to country, while maintaining adequate supplies of blood-derived products.
A risk assessment pertaining to a particular country should be undertaken in order to recommend appropriate risk-mitigating measures to employ in particular circumstances. Several countries, including Australia, Canada, France, Germany, UK, and the USA, have undertaken risk assessments using various modeling techniques and assumptions. Such risk assessments provide a basis for examining the adequacy of diff erent measures to minimize the risks to humans from human-derived materials and provide a framework for developing regulatory and public health actions. In some countries, risk assessments are carried out by expert groups independent from, but at the request of, the national regulatory agencies that recommend or select risk management actions. However, it is accepted that assessments cannot predict risk precisely, because of great uncertainties in the assumptions used. Risk assessments, while useful tools for developing risk-miti- gating strategies, are not the only tool. New information on vCJD prevalence and accumulation of actual data on transmission by blood will inevitably allow refi nement of risk evaluation. As new technologies, such as better testing for PrPTSE and practical procedures for reducing infectivity, are developed and validated, risk assessments can be further refi ned and risk-mitigating measures revised. Risk management should consider actions mitigating the consequences of both histori- cal risk and current risk, which may need diff erent approaches. Risk communication is another important component of policy but an aspect fraught with special diffi culties. It is important to decide on a signifi cant threshold of estimated risk—the threshold at which individuals are consid- ered to be at increased risk. No less important is to decide on an appropriate message, who should convey the message and how. Th e implications and practical consequences of notifying a group of people—most of whom are not expected to become ill—that they are at increased risk for vCJD must be carefully considered. Authorities should balance the public health goals of a notifi cation against the potential for causing individual harm—not the least of which is anxiety—imposed on individuals and their families, particularly because there is no practical action available to identify those persons who were actually infected or to help them avert illness.
Th e risk of secondary transmission of vCJD through blood, blood components and plasma derivatives depends upon the prevalence of vCJD in the donor population, both clinical cases and inapparent or pre-clinical infections, within a country. To consider appropriate measures to protect human and animal health, national authorities should seek two kinds of information regarding the risk of vCJD:
(a) Prevalence of BSE infection in native and imported cattle populations and potential BSE agent contamination of products (“internal” and “external” risk factors described below)
(b) Potential human exposures to the BSE agent
Th e minimal infectious doses of the BSE agent for humans exposed by various routes (the oral route presumed to be most common) are not known, and not every exposure is expected to result in infection. However, all potential opportunities for human exposure should be minimized.
Th e likelihood of human exposure to BSE depends upon internal and external factors:
— Internal factors. Th e internal or national risk of exposure depends on the geographical risk of BSE infections in cattle and the domestic patterns of preparing and using bovine-derived products: (i) cattle feeding practices—especially intentional or accidental use of feeds containing ruminant- derived non-milk proteins like meat-and-bone meal; (ii) slaughter practices, including age of cattle at slaughter, stunning techniques, removal of high-risk tissues from carcasses, containment and disposal of high-risk tissues; and (iii) the national use and distribution of meat and meat products.
— External factors. External risk is the potential exposure of humans to the BSE agent through the importation of infected animals or contaminated animal products and through exposure
13 of persons traveling outside the country in geographical areas where BSE is present in native or
imported beef products and where appropriate risk-reducing controls have not been implemented.
Th e joint meeting of WHO/FAO/OIE in 2001 (http://www.who.int/zoonoses/diseases/en) encouraged all countries to evaluate their potential exposure to BSE through systematic assessment of trade data and other possible risk factors. Such assessments are essential to identify and address risks to protect public health and prevent further spread of BSE.
In most cases the likelihood of vCJD in a country that has no BSE in cattle depends upon the extent to which people were either exposed outside the country or consumed imported commodities, such as beef products and by-products, contaminated with BSE agent. It is clear that countries without BSE in native cattle can still have cases of vCJD. Countries must be prepared to investigate vCJD cases with careful attention to possible internal and external exposures to the BSE agent.
Evidence from studies of blood from infected animals plus recent limited but convinc- ing evidence from human case reports, all indicate that blood transfusion effi ciently transmits some TSEs, including vCJD. Leukoreduction is known to remove some TSE infectivity from endogenously infected blood and might be expected to lower risk from labile blood components for transfusion without abolishing risk completely. Leukoreduction is already used widely in some countries, largely for other reasons. Unfortunately, recent studies found substantial endogenous infectivity remaining in plasma after blood of scrapie-infected hamsters was passed through a commercial leukoreduction fi lter.
For plasma-derived products, the major factors that infl uence overall risk—as estimated by sensitivity analysis in vCJD risk assessments—include the prevalence of vCJD, the estimated number of vCJD donations per plasma pool, and possible reduction of the TSE infectivity provided by some steps during manufacture. Th ere is a caveat for interpretation of investigational TSE clearance studies: because the natural physical form of the infectious TSE agent in blood is not known, the relevance of studies spiking various materials derived from infected brain tissue into blood that is then processed by scaled-down manufacturing steps remains unclear. Nevertheless, some national regulatory authorities have accepted the results of such studies—with all their uncertainties—as providing assurance that processes used to fractionate plasma are likely to reduce the risk from certain plasma derivatives. Th at conclusion has been reinforced by epidemiological studies in the UK, where, so far, no case of vCJD has been identifi ed in any recipient of plasma derivatives.
Risk-reducing measures
Product retrieval and market withdrawal
Cases of vCJD have been reported in persons who previously donated blood. When such cases are identifi ed, measures must be considered to reduce the risk of person-to-person transmission of infection, including the retrieval of in-date blood and blood components including plasma, as well as pools of plasma and manufacturing intermediates containing the donation. As a precaution, plasma derivatives prepared from those pools should also be identifi ed and withdrawn from the market. In countries with good systems for tracing recipients of blood components from donors later diagnosed with vCJD, the recipients should be notifi ed in an appropriate fashion and enrolled in follow-up surveillance. When a risk assessment indicates that the general population has been potentially exposed to BSE agent through food, additional actions to minimize risks should be considered, varying according to the probable prevalence of vCJD infections in the country.
Donor deferral
In countries where the risks of BSE in cattle and imports of commodities contaminated with BSE agent are both considered to be low or minimal, another possible source of vCJD infection remains—travel outside the country to areas where the BSE agent might have been present in meat products. In view of the probable long incubation periods of vCJD after oral exposure to the BSE agent—in excess of ten years—risk assessments must recognize that the travel to a country with
